Toxicological Characterization of Exon Skipping Phosphorodiamidate Morpholino Oligomers (PMOs) in Non-human Primates

2016 ◽  
Vol 3 (3) ◽  
pp. 381-393 ◽  
Author(s):  
Michael P. Carver ◽  
Jay S. Charleston ◽  
Courtney Shanks ◽  
Jianbo Zhang ◽  
Mark Mense ◽  
...  
Keyword(s):  
Exon Skipping ◽  
Phosphorodiamidate Morpholino Oligomers

scholarly journals Development of antisense-mediated myostatin knockdown for the treatment of insulin resistance

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Wouter Eilers ◽  
Mark Cleasby ◽  
Keith Foster
Keyword(s):  
Glucose Uptake ◽  
Muscle Mass ◽  
Exon Skipping ◽  
Negative Regulator ◽  
Intramuscular Administration ◽  
Transcript Levels ◽  
Muscle Disorders ◽  
Intravenous Injections ◽  
Phosphorodiamidate Morpholino Oligomers

AbstractMyostatin is a negative regulator of muscle mass and its inhibition represents a promising strategy for the treatment of muscle disorders and type 2 diabetes. However, there is currently no clinically effective myostatin inhibitor, and therefore novel methods are required. We evaluated the use of antisense phosphorodiamidate morpholino oligomers (PMO) to reduce myostatin expression in skeletal muscle and measured their effects on muscle mass and glucose uptake. C57/Bl6 mice received intramuscular or intravenous injections of anti-myostatin PMOs. Repeated intramuscular administration lead to a reduction in myostatin transcript levels (~ 20–40%), and an increase in muscle mass in chow and high-fat diet (HFD)-fed mice, but insulin-stimulated glucose uptake was reduced in PMO-treated muscles of HFD-fed mice. Five weekly intravenous administrations of 100 nmol PMO did not reduce myostatin expression, and therefore had no significant physiological effects. Unexpectedly, exon skipping levels were higher after intramuscular administration of PMO in HFD- than chow-fed mice. These results suggest that a modest PMO-induced reduction in myostatin transcript levels is sufficient to induce an increase in muscle mass, but that a greater degree of inhibition may be required to improve muscle glucose uptake.

Abstract 344: Finding the Achilles’ heel of Muscle Giant---TALEN-mediated Gene-editing in Zebrafish Titin

2015 ◽  
Vol 117 (suppl_1) ◽  
Author(s):  
Yu-Huan Shih ◽  
Xiaolei Xu
Keyword(s):  
Alternative Splicing ◽  
Gene Editing ◽  
Muscle Development ◽  
Exon Skipping ◽  
Transcription Activator ◽  
Detailed Characterization ◽  
A Domains ◽  
Underlying Mechanisms ◽  
Sarcomere Assembly

Background: TITIN (TTN) has more than 300 exons and encodes a gigantic protein that is crucial for heart and muscle development. Mutations in TTN caused a variety of human diseases including cardiomyopathy and muscular dystrophy. Recently, dilated cardiomyopathy-associated mutations on TTN have been found more frequently in exons encoding A-band domains but less in exons encoding the N-terminal Z-disc domains, suggesting that mutations in different exons of TTN cause distinct consequences. To elucidate the underlying mechanisms, we leveraged the Transcription Activator-Like Effects Nuclease (TALEN) technology in zebrafish to introduce truncating mutations in different exons of ttn, and then study their effects on heart and somites. Results: We generated truncational mutations in different exons of zebrafish titins encoding Z-disc, N2B, Novex-3, and A domains, respectively. Because zebrafish contains two titin homologues, ttna and ttnb, we introduced mutations in both genes at the corresponding loci. While both Z-disc and A band mutations on ttna disrupted sarcomere assembly in heart and somites, Z-disc or A band mutations on ttnb only affect somites without affecting the heart. Interestingly, a Z-disc mutation on ttna resulted in milder phenotypes than an A-band mutation, while a Z-disc mutation on ttnb generated severer phenotypes than an A-band mutation. No phenotype was observed in the homozygous fish in either ttna-novex-3 or ttnb-N2B mutant fish. Conclusions: A spectrum of truncational mutations in ttna and ttnb has been generated in zebrafish using the TALEN technology. Mutations in different exons result in different phenotypes. Detailed characterization of these mutants and double mutants will be presented, which shall elicit distinct contribution of alternative splicing and exon skipping as two candidate mechanisms during pathogenesis of Titinopathies.

Characterization of the human argininosuccinate lyase gene and analysis of exon skipping

Genomics ◽  
1991 ◽  
Vol 10 (1) ◽  
pp. 126-132 ◽  
Author(s):  
Richard D. Abramson ◽  
Peter Barbosa ◽  
Karen Kalumuck ◽  
William E. O'Brien
Keyword(s):  
Exon Skipping ◽  
Argininosuccinate Lyase

WED 237 Phosphorodiamidate morpholino oligomers for treatment of duchenne muscular dystrophy

2018 ◽  
Vol 89 (10) ◽  
pp. A34.2-A34
Author(s):  
Maresh Kate ◽  
Tiet May ◽  
Guglieri Michela ◽  
Domingos Joana ◽  
Straub Volker ◽  
...  
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Western Blot ◽  
Clinical Studies ◽  
Novel Mutation ◽  
Cumulative Risk ◽  
Exon Skipping ◽  
Dystrophin Expression ◽  
Polymerase Chain ◽  
Phosphorodiamidate Morpholino Oligomers

Exon skipping is a novel, mutation-specific approach to treating patients with Duchenne muscular dystrophy (DMD). Phosphorodiamidate morpholino oligomers are nucleic acid analogues that selectively redirect pre-mRNA splicing to enable production of internally truncated dystrophin.In exon 51 skipping (eteplirsen; n=36) and exon 53 skipping (golodirsen; n=25) clinical studies, internally shortened dystrophin mRNA was observed in all treated patients (per reverse transcription polymerase chain reaction). Eteplirsen increased dystrophin expression 15.5-fold, 11.6-fold, and 2.4-fold vs untreated controls (percent dystrophin-positive fibres, Western blot, and immunohistochemistry intensity, respectively; all, p≤0.007) in a 180 week study, and 2.8-fold (Western blot; p=0.008) in a 48 week study. Golodirsen increased dystrophin expression 10.7-fold (Western blot) over baseline following 48 weeks of treatment. Over 4 years, versus comparable external controls, eteplirsen slowed ambulatory decline (6 min walk test difference, 165 m; p=0.001) and cumulative risk of losing ambulation (83% vs 17%). In 2 clinical studies that included non-ambulatory patients, eteplirsen slowed pulmonary decline versus natural history data (assessed by spirometry).Eteplirsen and golodirsen demonstrated clinical and biochemical effects in patients with DMD; ongoing studies of these compounds are further characterising their effects in various patient populations.

scholarly journals Nonsequential Splicing Events Alter Antisense-Mediated Exon Skipping Outcome in COL7A1

2020 ◽  
Vol 21 (20) ◽  
pp. 7705
Author(s):  
Kristin A. Ham ◽  
May Thandar Aung-Htut ◽  
Sue Fletcher ◽  
Steve D. Wilton
Keyword(s):  
Intron Retention ◽  
Genetic Disorders ◽  
Exon Skipping ◽  
Monogenic Disease ◽  
Dystrophic Epidermolysis Bullosa ◽  
Mature Mrna ◽  
Intron Removal ◽  
Phosphorodiamidate Morpholino Oligomers ◽  
Splicing Patterns ◽  
Antisense Oligomer

The COL7A1 gene encodes homotrimer fibrils essential for anchoring dermal and epidermal layers, and pathogenic mutations in COL7A1 can cause recessive or dominant dystrophic epidermolysis bullosa. As a monogenic disease gene, COL7A1 constitutes a potential target for antisense oligomer-mediated exon skipping, a therapy applicable to a growing number of other genetic disorders. However, certain characteristics of COL7A1: many exons, low average intron size, and repetitive and guanine-cytosine rich coding sequence, present challenges to the design of specific and effective antisense oligomers. While targeting COL7A1 exons 10 and 73 for excision from the mature mRNA, we discovered that antisense oligomers comprised of 2′-O-methyl modified bases on a phosphorothioate backbone and phosphorodiamidate morpholino oligomers produced similar, but distinctive, splicing patterns including excision of adjacent nontargeted exons and/or retention of nearby introns in some transcripts. We found that the nonsequential splicing of certain introns may alter pre-mRNA processing during antisense oligomer-mediated exon skipping and, therefore, additional studies are required to determine if the order of intron removal influences multiexon skipping and/or intron retention in processing of the COL7A1 pre-mRNA.

scholarly journals Identification and Partial Characterization of a Variant of Human CXCR3 Generated by Posttranscriptional Exon Skipping

2004 ◽  
Vol 173 (10) ◽  
pp. 6234-6240 ◽  
Author(s):  
Jan Erik Ehlert ◽  
Christina A. Addison ◽  
Marie D. Burdick ◽  
Steven L. Kunkel ◽  
Robert M. Strieter
Keyword(s):  
Exon Skipping ◽  
Partial Characterization

scholarly journals New approaches to the treatment of orphan genetic disorders: Mitigating molecular pathologies using chemicals

2015 ◽  
Vol 87 (2 suppl) ◽  
pp. 1375-1388 ◽  
Author(s):  
RENATA V. VELHO ◽  
FERNANDA SPERB-LUDWIG ◽  
IDA V.D. SCHWARTZ
Keyword(s):  
Molecular Level ◽  
Genetic Disorders ◽  
Exon Skipping ◽  
Molecular Techniques ◽  
Genetic Mutations ◽  
Therapeutic Approaches ◽  
Translational Readthrough ◽  
Different Types ◽  
Identification And Characterization

With the advance and popularization of molecular techniques, the identification of genetic mutations that cause diseases has increased dramatically. Thus, the number of laboratories available to investigate a given disorder and the number of subsequent diagnosis have increased over time. Although it is necessary to identify mutations and provide diagnosis, it is also critical to develop specific therapeutic approaches based on this information. This review aims to highlight recent advances in mutation-targeted therapies with chemicals that mitigate mutational pathology at the molecular level, for disorders that, for the most part, have no effective treatment. Currently, there are several strategies being used to correct different types of mutations, including the following: the identification and characterization of translational readthrough compounds; antisense oligonucleotide-mediated splicing redirection; mismatch repair; and exon skipping. These therapies and other approaches are reviewed in this paper.

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